Lewis J K, Bothner B, Smith T J, Siuzdak G
Departments of Molecular Biology and Chemistry, The Scripps Research Institute, La Jolla, CA 92037, USA.
Proc Natl Acad Sci U S A. 1998 Jun 9;95(12):6774-8. doi: 10.1073/pnas.95.12.6774.
A dynamic capsid is critical to the events that shape the viral life cycle; events such as cell attachment, cell entry, and nucleic acid release demand a highly mobile viral surface. Protein mass mapping of the common cold virus, human rhinovirus 14 (HRV14), revealed both viral structural dynamics and the inhibition of such dynamics with an antiviral agent, WIN 52084. Viral capsid digestion fragments resulting from proteolytic time-course experiments provided structural information in good agreement with the HRV14 three-dimensional crystal structure. As expected, initial digestion fragments included peptides from the capsid protein VP1. This observation was expected because VP1 is the most external viral protein. Initial digestion fragments also included peptides belonging to VP4, the most internal capsid protein. The mass spectral results together with x-ray crystallography data provide information consistent with a "breathing" model of the viral capsid. Whereas the crystal structure of HRV14 shows VP4 to be the most internal capsid protein, mass spectral results show VP4 fragments to be among the first digestion fragments observed. Taken together this information demonstrates that VP4 is transiently exposed to the viral surface via viral breathing. Comparative digests of HRV14 in the presence and absence of WIN 52084 revealed a dramatic inhibition of digestion. These results indicate that the binding of the antiviral agent not only causes local conformational changes in the drug binding pocket but actually stabilizes the entire viral capsid against enzymatic degradation. Viral capsid mass mapping provides a fast and sensitive method for probing viral structural dynamics as well as providing a means for investigating antiviral drug efficacy.
动态衣壳对于塑造病毒生命周期的事件至关重要；诸如细胞附着、细胞进入和核酸释放等事件需要高度可移动的病毒表面。普通感冒病毒人鼻病毒14型（HRV14）的蛋白质质量图谱揭示了病毒的结构动力学以及抗病毒药物WIN 52084对这种动力学的抑制作用。蛋白水解时间进程实验产生的病毒衣壳消化片段提供了与HRV14三维晶体结构高度一致的结构信息。正如预期的那样，初始消化片段包括衣壳蛋白VP1的肽段。这一观察结果在意料之中，因为VP1是最外层的病毒蛋白。初始消化片段还包括属于VP4的肽段，VP4是最内层的衣壳蛋白。质谱结果与X射线晶体学数据共同提供了与病毒衣壳“呼吸”模型一致的信息。虽然HRV14的晶体结构显示VP4是最内层的衣壳蛋白，但质谱结果显示VP4片段是最早观察到的消化片段之一。综合这些信息表明，VP4通过病毒呼吸短暂暴露于病毒表面。在有和没有WIN 52084的情况下对HRV14进行的比较消化显示消化受到显著抑制。这些结果表明，抗病毒药物的结合不仅会导致药物结合口袋处的局部构象变化，实际上还会稳定整个病毒衣壳以抵抗酶促降解。病毒衣壳质量图谱提供了一种快速且灵敏的方法来探测病毒结构动力学，同时也为研究抗病毒药物疗效提供了一种手段。
